BODIPY-Au(I): A Photosensitizer for Singlet Oxygen Generation and Photodynamic Therapy

Article abstract of DOI:10.1021/acs.orglett.7b00791

Upon complexation with Au(I), a photoinactive BODIPY derivative was transformed into a highly photoactive triplet sensitizer. Along with high efficiency in singlet oxygen generation (φ_δ = 0.84), the new BODIPY-Au(I) skeleton showed excellent ph

Full text of DOI:10.1021/acs.orglett.7b00791

Letter
pubs.acs.org/OrgLett
BODIPY−Au(I): A Photosensitizer for Singlet Oxygen Generation and
Photodynamic Therapy
‡
‡
̈
Muhammed Ucu
̧
̈
ncu
̈
, Erman Karakus,
̧ Eylem Kurulgan Demirci, Melike Sayar, Suay Dartar,
and Mustafa Emrullahogl
̆
u*
̇
Department of Chemistry, Faculty of Science, Izmir Institute of Technology, Urla, 35430 Izmir, Turkey
*
S Supporting Information
ABSTRACT: Upon complexation with Au(I), a photoinactive BODIPY
derivative was transformed into a highly photoactive triplet sensitizer. Along
with high efficiency in singlet oxygen generation (Φ = 0.84), the new
Δ
BODIPY−Au(I) skeleton showed excellent photocytotoxic activity against
cancer cell lines (EC = 2.5 nM).
5
0
he search for new classes of photosensitizers with
promote spin−orbit coupling, which allows the sort of
T
potential applications in photodynamic therapy (PDT)
intersystem crossing (ISC) needed to observe triplet states
1
has intensified significantly in the past few years. A well-
(
Scheme 1a).
2
recognized clinical modality for treating localized cancers and
3
1
other diseases, PDT involves the activation of a photo-
Scheme 1. BODIPY-Based Photosensitizers for O
Generation
2
sensitizer by light to generate cytotoxic reactive oxygen species
e.g., singlet oxygen) within cells to promote irreversible
(
cellular damage and cell death. The critical component of PDT
is the photosensitizer, which should have a high capacity for
singlet oxygen generation, absorption bands within the
therapeutic window, and the ability to colocalize in cancer
tissue.
Most photosensitizers investigated in clinical trials comprise
cyclic tetrapyrrole structures inspired by naturally derived
1d,4
porphyrins such as hematoporphyrin.
Although synthetic
dyes not of porphyrin origin have also been evaluated for their
5
photosensitizing abilities, they suffer from drawbacks such as
dark cytotoxicity and photobleaching.
As an alternative class to nonporphyrin-based photo-
sensitizers, the boron−dipyrromethene (BODIPY) dye plat-
6
As alternatives to heavy atom-bearing BODIPY dyes,
orthogonal BODIPY dimers and trimers have been reported
by Akkaya et al. and Zhang et al. to be exceptionally
form has also been subject to extensive study. BODIPY dyes
have certain characteristics, including high extinction coef-
ficients, robustness toward light and chemicals, and resistance
to photobleaching, that make them good candidates as
14
15
effective photosensitizers for singlet oxygen generation
7
(
Scheme 1b). In contrast, Zhao et al. have identified
photosensitizers for PDT. However, deprived of excited triplet
BODIPY−C as a heavy atom-free organic triplet sensitizer
states, which represent a key photophysical parameter necessary
for generating singlet oxygen from molecular oxygen, the
unmodified BODIPY core (e.g., tetramethyl-BODIPY) remains
photoinactive.
60
16
that uses C as a spin converter (Scheme 1c).
60
Introducing transition-metal ions in close proximity to a
fluorophore skeleton is another approach to enabling the
observation of excited triplet states. In that context, a range of
transition-metal complexes of BODIPY-based luminophores
have been designed and investigated as triplet photosensi-
Recent studies on BODIPY-based photosensitizers have
elegantly demonstrated that the controlled manipulation of the
BODIPY core could precipitate intriguing photophysical
17−21
8
9
10
11
tizers.
Nevertheless, to the best of our knowledge, using a
changes. In that context, Nagano, Akkaya, O’Shea, and
1
2
13
McClenaghan, as well as other researchers, have clearly
established that placing heavy atoms (e.g., bromine or iodine)
at appropriate positions on the primary BODIPY core can
Received: March 16, 2017
©
XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.7b00791
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
BODIPY−Au(I) complex as a singlet oxygen generator has yet
to be examined.
To develop a new chemical construct as an efficient singlet
oxygen generator, we designed a BODIPY dye platform
comprising −LAu(I) (L = PPh ) as a potential spin converter
3
(
S −T ) (Scheme 1d). We anticipated that incorporating
1
1
−
LAu(I) into the alkynyl backbone of a BODIPY core would
enhance the probability of ISC due to the heavy atom effect of
Au(I) and thereby improve the possibility to observe excited
triplet states.
Herein, we present the design, synthesis, and spectroscopic
investigation of a BODIPY−Au(I) construct, denoted as
BOD−Au, which allows for exceptionally rapid, highly efficient
singlet oxygen generation both in solution and living
environments. We also established the construct’s great
potential for use as a cytotoxic agent in PDT.
We prepared BOD−Au synthetically, as outlined in Scheme
. We treated the individually prepared acetylene derivative of
2
Scheme 2. Synthesis of BOD-Au
Figure 1. (a) Absorption spectra of BOD-Ac (5 μM) and BOD-Au (5
μM) in CH Cl ; (b) emission spectra of BOD-Ac (5 μM) and BOD-
BODIPY (BOD−Ac) in methanol with AuClPPh3 in the
presence of a base (i.e., NaOH) to generate the compound in a
reasonable yield of approximately 75%. We adopted recrystal-
lization as the protocol for purification since we observed
2
2
Au (5 μM) in CH Cl (λ : 460 nm for BOD-Ac, λ : 540 nm for
2
2
ex
ex
BOD-Au).
decomposition during chromatography on SiO . We confirmed
2
the identity of the title compound via nuclear magnetic
resonance spectroscopy and high-resolution mass spectrometric
and briefer fluorescence lifetime than that of BOD−Ac (Φ
=
F
0.58, τ /(ns) = 5.38), as Table 1 shows.
F
2
2
analysis.
Although a detailed investigation of the excited-state
properties of BOD−Au remains necessary (e.g., via transient
absorption spectroscopy), a sharp decrease in Φ and briefer
F
We investigated the spectroscopic behavior of BOD−Au and
its ability to generate singlet oxygen by fluorescence and
absorption spectroscopy. Table 1 shows the electronic
absorption and emission data of BOD−Au and, for
comparison, of the gold-ion-free derivative BOD−Ac.
decay time of the singlet excited state indicated ISC promoted
by the internal heavy atom effect of Au(I). Importantly, BOD−
Au displayed no phosphorescence at either room temperature
or 77 K. Similar results for some transition-metal-ion complexes
of BODIPY dyes have been observed by other researchers as
Table 1. Photophysical Parameters of BOD-Ac, BOD-Au,
and 2I-BOD
21,24
well.
To confirm that BODIPY can generate T states via ISC, a
1
a
b
c
d
e
compd
λabs
ε
λem
ΦF
τF (ns)
ΦΔ
reasonable approach is to use molecular oxygen as a triplet
acceptor, which in turn generates singlet oxygen via triplet−
triplet state energy transfer. To that end, we assessed the
capabilities of BOD−Au and BOD−Ac to generate singlet
g
BOD-Ac
BOD-Au
518
550
529
5.1
3.8
8.5
535
583
548
0.58
0.05
0.03
5.38
1.02
na
0.84
f
2
I-BOD
0.13
0.79
a
In CH Cl2 (5 × 10−6 M).4 Molar absorption coefficient at the
b
oxygen by employing a trapping method using diphenyliso-
2
−1
−1
c
1
absorption maximum, ε: 10
M
cm . With rhodamine B as
benzofuran (DPBF) to trap O . Singlet oxygen interacts with
2
d
e
standard (Φ = 0.31 in H O). Fluorescence lifetimes Singlet oxygen
DPBF to yield 1,2-dibenzoylbenzene, and the extent of DPBF-
related photodegradation can be evaluated by measuring the
decrease in the DPBF absorption band at 415 nm (Figure 2).
For BOD−Ac, we detected no signs of DPBF photo-
degradation, whereas for BOD−Au, absorbance at 415 nm
decreased remarkably and disappeared entirely within a couple
F
2
1
f
23 g
(
O ) quantum yield. In CH CN. Not applicable.
2 3
As Figure 1 reveals, the absorption spectra of BOD−Ac is
dominated by a maximum band at approximately 520 nm
belonging to spin-allowed π−π* transitions of the BODIPY
core. For BOD−Au, the absorption band was significantly red-
shifted compared to its metal-free analogue, which clearly
indicated a perturbation on the π-conjugation. Upon excitation
at 540 nm (CH Cl , 5 μM), BOD−Au also exhibited a red-
of minutes, which clearly demonstrates its high efficiency in
1
generating O . The efficiency of generating singlet oxygen,
2
known as singlet oxygen quantum yield, has a remarkably high
Φ
value (Table 1), calculated with diiodo-BODIPY (2I−
Δ
23 22
BOD ) as the reference. Our findings show that BOD−Au is
one of the most efficient BODIPY-based singlet oxygen
generator developed thus far. Meanwhile, the absorbance of
2
2
shifted emission band in the visible region, albeit with a far
smaller fluorescence quantum yield (Φ = 0.05, τ /(ns) = 1.02)
F
F
B
DOI: 10.1021/acs.orglett.7b00791
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
Figure 3. Fluorescence images of human lung adenocarcinoma cells
(
A549). (a) Bright field image of A549 cells treated with only BOD-
Au (10 μM); (b) fluorescence image of cells treated with Lyso-tracker
control); (c) fluorescence image of cells treated with BOD-Au (10
(
μM); (d) merged images of a−c.
Figure 2. (a) Singlet oxygen mediated bleaching of DPBF in the
presence of BOD-Au (5 μM in CH Cl ). (b) Comparison of the rates
2
2
of decay of DPBF (25 μM) in CH Cl , as monitored at 415 nm, using
2
2
BOD-Ac, BOD-Au as the photosensitizers (5 μM) and 2I-BOD (5
2
μM) as the reference. Irradiation using green LED (3.3 mW/cm )
emitting at λem = 525 nm at the distance of 15 cm from the cuvette
window.
Figure 4. (a) Cell viability of A549 cells after treatment with BOD−
Au at different concentrations (2.5, 5, 10, and 20 nM). Control group
was incubated only with the cell culture medium. (b) Effect of light
dose on cell viability (BOD−-Au, 20 nM).
BOD-Au at 550 nm remained stable during the irradiating
process, thus proving its high resistance to photobleaching
(
Figure 2a).
The promising efficiency of BOD−Au in generating singlet
oxygen in the solution encouraged us to further assess its in
vitro photocytotoxic activity against cancer cell lines. To
ascertain the subcellular localization of BOD−Au within A549
cells and observe whether any correlation exists with MTT
assay results, we first studied its subcellular localization within
A549 cells by using LysoTracker as an organelle-specific
fluorescent dye to stain the lysosomes. Although the
fluorescence emission of BOD−Au was relatively weak in the
solution, it was sufficient for its visualization in the cellular
medium. Based on the counter stain and green fluorescence
emitting from the cells, we thus conclude that the BOD−Au
passes through the cell membrane and localizes particularly in
the cytosol (Figure 3).
For cell lines kept in the dark, however, BOD−Au showed no
significant cytotoxicity at concentrations up to 40 nM (IC
=
5
0
22
0
.16 μM). We also examined the change in cell viability as a
function of the light dosage; results clearly showed that the
extent of light illumination is critical to the photodynamic
efficiency of the photosensitizer.
In summary, we devised a photosensitizer comprising a
BODIPY dye as a visible light-harvesting chromophore and
−
LAu(I) (L = PPh ) as a spin convertor (S −T ). By
3
1
1
introducing −LAu(I) into the backbone of a BODIPY dye,
we transformed a photoinactive dye into a highly photoactive
triplet sensitizer. In line with its high singlet oxygen generation
efficiency (Φ = 0.84), this new BODIPY-based photo-
Δ
The photodynamic activity of BOD−Au in Tween 80
sensitizer showed excellent photocytotoxic activity against
cancer cell lines (EC50 = 2.5 nM).
emulsions was investigated against A549 cells by using MTT
2
2
assay. We first loaded A549 cells with BOD−Au, irradiated
2
them with green light (green LED, 525 nm, 3.3 mW/cm ) for
ASSOCIATED CONTENT
Supporting Information
■
5
−30 min, and incubated them for 48 h in the dark. We
*
S
protected another group of A549 cells from light throughout
the incubation process in order to explore the effect of light on
the activity of BOD−Au.
For cell lines illuminated by green light, we observed a
significant decrease in viability even at very low concentrations
of BOD−Au (EC = 2.5 nM), as the red bars in Figure 4 show.
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.7b00791.
Absorbance, fluorescence, and characterization data and
experimental procedures (PDF)
5
0
C
DOI: 10.1021/acs.orglett.7b00791
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
A.; Killoran, J.; O’Shea, C.; Kenna, T.; Gallagher, W. M.; O’Shea, D. F.
J. Am. Chem. Soc. 2004, 126, 10619−10631.
AUTHOR INFORMATION
■
Corresponding Author
(
12) Batat, P.; Cantuel, M.; Jonusauskas, G.; Scarpantonio, L.; Palma,
*
E-mail: mustafaemrullahoglu@iyte.edu.tr.
A.; O’Shea, D. F.; McClenaghan, N. D. J. Phys. Chem. A 2011, 115,
4034−14039.
13) Adarsh, N.; Avirah, R. R.; Ramaiah, D. Org. Lett. 2010, 12,
5720−5723.
14) (a) Cakmak, Y.; Kolemen, S.; Duman, S.; Dede, Y.; Dolen, Y.;
Kilic, B.; Kostereli, Z.; Yildirim, L. T.; Dogan, A. L.; Guc, D.; Akkaya,
E. U. Angew. Chem., Int. Ed. 2011, 50, 11937−11941. (b) Duman, S.;
Cakmak, Y.; Kolemen, S.; Akkaya, E. U.; Dede, Y. J. Org. Chem. 2012,
1
(
ORCID
̆
Mustafa Emrullahoglu: 0000-0002-8221-2597
Author Contributions
(
‡
̈
M.U. and E.K. contributed equally.
Notes
7
7, 4516−4527. (c) Kolemen, S.; Is ı̧ k, M.; Kim, G. M.; Kim, D.; Geng,
The authors declare no competing financial interest.
H.; Buyuktemiz, M.; Karatas, T.; Zhang, X.-F.; Dede, Y.; Yoon, J.;
Akkaya, E. U. Angew. Chem., Int. Ed. 2015, 54, 5340−5344.
(d) Ozdemir, T.; Bila, J. L.; Sozmen, F.; Yildirim, L. T.; Akkaya, E.
U. Org. Lett. 2016, 18, 4821−4823.
ACKNOWLEDGMENTS
■
̇
We thank Izmir Institute of Technology (IZTECH) and
Turkish Academy of Science, Outstanding Young Scientist
(
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5
539. (b) Pang, W.; Zhang, X.-F.; Zhou, J.; Yu, C.; Hao, E.; Jiao, L.
̇
̇
Award (TUBA-GEBIP-2016), for financial support and Izmir
Institute of Technology, Biotechnology and Bioengineering
Research and Application Centre for fluorescence imaging
facilities.
Chem. Commun. 2012, 48, 5437−5439. (c) Zhang, X.-F.; Yang, X. J.
Phys. Chem. B 2013, 117, 9050−9055.
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DOI: 10.1021/acs.orglett.7b00791
Org. Lett. XXXX, XXX, XXX−XXX